In the last decades, epidemiological long term studies have identified numerous factors accelerating formation of atherosclerosis and thereby promoting development of cardiac infarctions. Despite of avoiding risk factors and of behavior increasing the risk of atherosclerosis, the development of pronounced atherosclerotic changes culminating in heart infarction can even be observed in young adults. In such cases, genetic factors play a decisive role. It is for example known that defects of genes playing an important role in cholesterol metabolism require medication with cholesterol lowering drugs. The early detection of a genetic defect allows that counter measures can be taken in time.
Therefore, it is desirable both from a diagnostic point of view and from a therapeutical point of view to be able to recognize and to detect crucial genetic alterations (see A. R. Miserez, Die Bedeutung genetischer Faktoren bei der Entstehung des Herzinfarkts, uni nova, April 1998, S. 44–52).
Cholesterol, besides being the precursor of steroid hormones and bile acids, is an essential constituent of the cell membrane decisively enhancing its permeability-barrier properties. Human cells control their intracellular cholesterol concentration tightly by regulating the receptor-mediated uptake of extracellular cholesterol-containing low density lipoproteins (LDL) and the intracellular cholesterol biosynthesis. LDL particles bind to the LDL receptor (LDLR) by their apolipoprotein (apo) B moieties. The binding and subsequent internalization of these lipoprotein-receptor complexes can be partially or completely abolished if one of the proteins involved in this process is defective or missing. Mutations of the genes encoding the apolipoprotein E (causing familial dysbetalipoproteinemia (FDL)), the apo B-100 (causing familial defective apo B (FDB)), and the LDL receptor (causing familial hypercholesterolemia (FH)) lead to an accumulation of cholesterol-containing particles in the plasma, which is associated with an increased risk of coronary artery disease. In most of the tested populations, said mutations can only explain 4.2 to 7% of cases with hypercholesterolemia (defined as the 10% of persons of a population with LDLC concentrations above the ninetieth percentile). Thus, the casual gene defects for the majority of affected people with increased plasma LDLC are not yet identified.
The promoters of the LDLR gene and of the genes involved in the cholesterol biosynthesis including the hydroxymethylglutaryl (HMG) CoA synthase, farnesyl-pyrophosphate synthase, and squalene synthase genes, contain specific nucleotide sequences, so-called sterol regulatory elements (SREs).
It is already known that two proteins, SRE-binding protein-(SREBP-) 1 and SREBP-2, bind the SREs in the promoters of these genes and activate their transcription rates. When cells are deprived of sterols, both proteins are activated by two proteolytic steps, first by a sterol-sensitive, and then by a cholesterol-independent step. These cleavage events release 68 kDa peptides from the NH2-terminal region of the SREBP-1 and -2 precursor proteins in the cytoplasm. The NH2-terminal, mature form of the transcription factors enters the nucleus and binds the SREs in the promoters of cholesterol-regulating genes. As a consequence, these genes are activated, thus leading to an increase in the receptor-mediated uptake of LDL as well as to an enhanced intracellular cholesterol biosynthesis.
When cholesterol accumulates in the cell, the first, cholesterol-sensitive cleavage event is inhibited, the mature forms of the SREBPs disappear and transcription rates decline, thereby preventing excessive accumulation of cholesterol in the cell. SREBP-1 and SREBP-2 regulate numerous SRE-containing genes involved in cholesterol homeostasis. In addition, SREBP-1 activates the HMG CoA reductase and the squalene synthase. SREBP-1 and SREBP-2 are members of the so-called basic helix-loop-helix leucine zipper transcription factor family. The genes encoding these factors have been cloned recently, and their genetic structures have been characterized (20,21).
Despite of the available knowledge, the percentage—as mentioned above—of identifiable risk patients for e.g. hypercholesterolemia is below 7%.
Therefore, the present invention had the aim to improve the early diagnosis and therapy of risk patients.
Said aim is achieved by providing diagnostic methods as well as polymorphisms in the SREBP genes which are suitable for the use in said diagnostic methods, in particular polymorphisms which are found in a fraction of patients with altered lipid metabolism, in particular cholesterol metabolism, preferably in a big fraction of such patients.